The overall goal of this PROGRAM PROJECT is to determine whether B7-1 and B7-2 costimulatory molecules provide the second signal which can induce either the afferent or efferent phases of autoreactive T cell activation leading to pathologic autoimmune disease. To this end, three projects have been constructed. In Project 1, mice lacking B7- and B7-2 molecules will be used to evaluate the potential importance of these costimulatory molecules on the incidence and severity of experimental autoimmune disease. These mice will provide unique models to determine whether costimulatory molecules are capable of providing the minimal second signals to drive autoreactive T cells to induce pathologic autoimmune disease. The central goal of Project 2 is to identify mechanisms by which altered peptide ligands (APLs) and B7 costimulatory molecules affect cytokine balance and suppress antigen-specific proinflammatory activity and enhance anti- inflammatory functions of T cells. The contribution of strength of signal/stimulus to T cell differentiation provided by B7 molecules an APLs will be examined using a panel of CHO transfectants with varying levels of B7 and class II MHC expression and a panel of APLs. Project 3 examines the role of costimulatory pathways in MS. The requirements for expression of costimulatory molecules and il-12 by human glial cells will be examined. The molecular basis for the activated in vivo state of autoreactive T cells in MS patients will be further investigated by comparing myelin antigen- autoreactive peripheral blood T cells in MS patients to controls and determining their capacity to expand in response to antigen presented by CHO cells transfected with MHC class II, DR2, alone or with B7-1 or B7-2 costimulatory molecules and DR2. Additionally, the role of costimulation in negative regulation of T cells will be examined. This Program exemplifies the most fundamental concept of a BASIC/CLINICAL PROGRAM PROJECT since all three projects are highly interdependent. It is through the interrelationship between projects that we wish to rapidly transfer technology from the bench to the understanding and therapy of human autoimmune disease. GRANT-P01AI396710001 Signaling via the B7-CD28/CTLA4 pathway can provide a potent costimulatory signal for T cell activation. For this reason, methods directed toward blocking this pathway have received considerable attention for the treatment of autoimmune disease. B7-CD28/CTLA4 pathway blockade may enable a distinctive treatment strategy, since it would affect only those antigen- specific T cells undergoing activation and not be globally immunosuppressive. We have recently found that murine CTLA4-Ig, which binds to both B7-1 and B7-2, completely inhibits the induction of experimental autoimmune encephalomyelitis (EAE) and induces long-lived protection. We will determine the cellular immunologic basis for the potent therapeutic effects of CTLA-4Ig, and compare it with anti-B7 mAb- mediated immunomodulation. However, the presence of multiple costimulatory molecules in the pathway together with the potential for both positive and negative signaling through the pathway indicate that additional insights are needed in order to successfully manipulate this key immunoregulatory pathway. Using transgenic approaches, we will focus in this project on the function of the B7-CD28/CTLA4 pathway in both the afferent arm of the autoimmune respones where T cells become activated in the periphery and in the effector arm where potentially pathogenic T cells are driven to mediate tissue destruction: Mice lacking B7-1 and/or B7-2 will be used to evaluate the importance of the B7 costimulatory molecules in the induction, reactivation and CNS pathologic stages of EAE. These unique tools will permit us to determine whether the elimination of B7-1 and/or B7-2 will block or modulate the induction and/or effector stages of EAE. These murine models allow us to test the biological significance of the observation of Project 3, that early MS plaques ia the CNS are associated with increased expression of B7-1. We will compare the induction and course of EAE in these knockouts, and in mice given specific B7 antagonists. We will analyze the mechanism of observed effects through the generation and analysis of T cell clones. With Project 2, we will investigate the epitope specificity of T cell clones generated from CTLA- 4Ig treated mice and B7 deficient mice, as a means to determine if there is a contribution of epitope shift to a protective phenotype. Taken together, these studies will define the role of B7-1 and B7-2 in autoimmune disease, enabling rational pharmacological manipulation of this important immunoregulatory pathway.